9  Differences in microglia by sample & treatment

9.1 Total microglia states

Uniform Manifold Approximation and Projection (UMAP) visualization of 29854 microglial cells.

Each point represents a single cell, and cells are colored according to unsupervised clusters identified from transcriptomic profiles.

9.2 Total cells

Bar plot showing the number of cells assigned to each cluster.

9.3 Gene Heatmap & hierarchical cluster

Heatmap of normalized expression levels for representative marker genes across clusters, with hierarchical clustering applied to both genes and clusters, providing an overview of the molecular signatures that distinguish microglial states.

9.4 Proportions (%) of Microglial States Across Samples and Treatments

Stacked barplot of the relative abundance of each microglial Seurat-defined cluster in individual samples and compared proportions between the Aducanumab-treated and IgG control groups.

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	Adu	IgG	p.overall
	N=3	N=3
NFkB/IEG Activated	23.3 [22.0;23.8]	25.9 [24.8;26.1]	0.127
Homeostatic (Activation competent)	14.9 [14.9;15.1]	19.6 [18.5;20.1]	0.050
DAM (Early/Intermediate)	18.5 [17.4;19.0]	13.9 [13.3;14.4]	0.050
Remodeling / Injury response	12.5 [12.3;12.6]	11.1 [11.1;11.8]	0.268
Homeostatic (canonical)	10.6 [10.4;11.8]	11.1 [10.2;12.6]	0.827
DAM (late)	11.2 [9.80;12.3]	11.8 [11.5;11.9]	0.658
Antigen presenting	7.10 [4.30;7.10]	2.40 [2.40;4.20]	0.500
Interferon responsive	3.60 [3.25;3.70]	2.70 [2.65;2.90]	0.127
Proliferating	0.60 [0.45;0.80]	0.50 [0.45;0.55]	0.658

Values are median % of microglia in that cluster per sample, with IQR.
p.overall: Mann-Whitney p-value

9.5 Module scores

UMAP displaying the combined expression scores or the distribution of individual canonical microglial marker genes, illustrating the transcriptional features that define each cluster.

Markers from bibliography to generate the scores

Signature	genes
AP	Cd74, B2m, Cd37, Lgals1, Rpl26, Rps19, Rps8, Rps4x, Rps15a, H2-Eb1, H2-Ab1, H2-Aa
CRM	Il1b, Ch25h, Padi2, Plek, Tlr2, Junb, C3ar1, Nfkbia, Trem2, Tyrobp
DAM	Spp1, Lipa, Apoc1, Ccl3, Fabp5, Trem2, Cxcr4, Gpnmb, Litaf, Cstb, Rps18, Rpl11, Rpl37a, Axl, Dbi
Homeostatic	P2ry13, P2ry12, Cx3cr1, Cst3, Ltc4s, Bin1, Maf, Frmd4a, Csf1r, C1qa, Dock8, Stab1, Sparc
IRM	Ifit2, Il1b, Epsti1, Eif2ak2, Bst2, Tnfsf10, Sp110, Irf1, Tap1, Adar, Cd48, Nampt
TIM	Klf4, Malat1, Tmx4, Jund, Btg2, Jun, Fos, Klf2

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Red dots indicate cells with high signature scores (top 25%), while grey represents cells with lower scores

Homeostatic
DAM: Disease associated microglia
AP : Antigen Presenting (HLA/MHC class II)
IRM : Interferon-response microglia.
CRM : Cytokine-Response microglia Markers described in (Fumagalli et al. 2025)

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TIM : Terminally inflammatory microglia (Millet, Ledo, and Tavazoie 2024)

Fumagalli, Laura, Alma Nazlie Mohebiany, Jessie Premereur, Paula Polanco Miquel, Baukje Bijnens, Pieter Van de Walle, Nicola Fattorelli, and Renzo Mancuso. 2025. “Microglia Heterogeneity, Modeling and Cell-State Annotation in Development and Neurodegeneration.” Nature Neuroscience 28 (7): 1381–92. https://doi.org/10.1038/s41593-025-01931-4.

Millet, Alon, Jose Henrique Ledo, and Sohail F. Tavazoie. 2024. “An Exhausted-Like Microglial Population Accumulates in Aged and APOE4 Genotype Alzheimer’s Brains.” Immunity 57 (1): 153–170.e6. https://doi.org/10.1016/j.immuni.2023.12.001.